1. Field of the Invention
The present invention relates to bacteria and bacterial combinations which can be used in methods to improve the health and vigor, including enhancement of the growth of plants, including important crop plants, while improving the sustainability of the agro-ecosystem. The bacterial strains herein include Paenibacillus sp. (ATY16); Bacillus megaterium (PT6); Bacillus subtilis (PT26A); and combinations thereof, and can be useful for treatment of healthy plants and plants which are susceptible to plant disease or which have been infected with plant disease. Although the methods and compositions are useful for administration to any plant or seed, preferred plants are those which are commercial crops, for example citrus, corn, soybean, and tomato. The methods and compositions of embodiments of the invention can ameliorate the effects of plant diseases, including microbial diseases such as huanglongbing (HLB) disease (also known as citrus greening disease).
2. Description of the Related Art
Conventional pest control technologies based on the use of agricultural chemicals have contributed to efficient agricultural productivity. However, their use also has led to increasing public concerns regarding their negative impacts on the environment. Environmentally-beneficial agriculture using no or reduced amounts of agricultural chemicals and satisfying cultivation efficiency, while assuring human safety is desired and necessary. Therefore, pest and disease control technology fulfilling such demand is needed in the art.
Crops in different ecosystems around the world may suffer less than ideal conditions due to soil or weather conditions, or various stresses, as well as diseases that can negatively affect the health and vigor of the crop plants. Such factors can reduce productivity of the crops to a greater or lesser degree, even under good growing conditions. Thus, crop plants can benefit from treatment that will increase the health and vigor of the plants, whether the plants are stressed by poor conditions, by disease, or even when the plants are healthy or grown under favorable conditions.
A number of plant diseases have negative effects on crop plants worldwide. Microbial plant pathogens can lead to losses in yield, and can even kill crop plants. Therefore, strategies to improve plant defenses against pathogens are needed to improve cultivation, crop yield, and crop quality, while avoiding environmental pollution of the plants and the soil in which they are grown. Biological approaches, such as the use of beneficial bacteria as described herein, therefore are helpful to improve crop plant health generally, and to reduce the effects of plant pathogens.
An example of a harmful plant disease is HLB or citrus greening disease also sometimes referred to as yellow shoot or yellow dragon. This is a major bacterial disease of citrus crops and can be found in Asia, in the Americas and in Africa. It has been spreading worldwide, resulting in economic loss. Huanglongbing (HLB) is currently the most economically devastating disease of citrus worldwide and no established cure is available. All commercial citrus varieties currently available are susceptible to HLB and the citrus industries in affected areas have suffered a decline in both production and profit (Bové, 2006; Gottwald et al., 2007; Wang and Trivedi, 2013). In Florida, HLB is now present in all commercial citrus-producing counties and is destroying the $9 billion citrus industry at a rapid pace. It was estimated that HLB has played a key role in the loss of about 100,000 citrus acres since 2007 in Florida and has cost Florida's economy approximately $3.6 billion in lost revenues since 2006 (Gottwald, 2010; Wang and Trivedi, 2013).
Citrus HLB is associated with a phloem-limited fastidious α-proteobacterium belonging to the ‘Candidatus’ genus Liberibacter, formerly known as Liberobacter (Jagoueix et al., 1994). Currently, three species of ‘Ca. Liberibacter’ have been identified to cause HLB disease: ‘Ca. L. asiaticus’ (Las), ‘Ca. L. africanus’, and ‘Ca. L. americanus’ (Gottwald, 2010). These bacteria have not been cultivated in pure culture. HLB pathogen is mainly spread by the insect (psyllid) vector Diaphorina citri in the field. There are two psyllid species transmitting Liberibacters: Asian citrus psyllid (Diaphorina citri) in Asia and the Americas (Bové, 2006; Halbert, 2005; Teixeira et al., 2005) and African citrus psyllid (Trioza erytreae) in Africa (Bové, 2006). Las and Asian citrus psyllid are the most prevalent and important throughout HLB-affected citrus-growing areas worldwide (Bové, 2006). Las propagates in the phloem of the host plants, resulting in die-back, small leaves, yellow shoots, blotchy mottles on leaves, corky veins, malformed and discolored fruit, aborted seed, premature fruit drop, root loss, and eventually tree death (Bové, 2006; Gottwald et al., 2007; Wang and Trivedi, 2013). The life span for the profitable productivity of infected citrus trees is dramatically shortened as the disease severity increases and the yield is significantly reduced while the tree is still alive (Gottwald et al., 2007). The understanding of virulence mechanism of the bacterial pathogen is limited, due to the difficulty in culturing Las. So far, most molecular insights of the HLB biology and Las pathogenicity are derived from the genome sequences of Las and other related Liberibacters (Duan et al., 2009; Lin et al., 2011; Leonard et al., 2012; Wulff et al., 2014).
Particularly sensitive citrus includes Citrus halimii, ‘Nules’ clementine mandarin, Valencia sweet orange, ‘Madam Vinous’ sweet orange, ‘Duncan’ grapefruit, ‘Ruby’ red grapefruit, and ‘Minneola’ tangelo, however, any Citrus species is vulnerable to HLB. In addition, some related plants in the genus Rutaceae, and other plants may become infected with Ca. Liberibacter species. Those of skill in the art are able to test for infection by Ca. Liberibacter, and therefore are able to determine which plants suffer from HLB or Ca. Liberibacter infection. Treatment of such plants is considered part of this invention.
Current methods in use for HLB control include the use of HLB-free citrus seedlings, destruction of infected trees, and application of insecticides such as aldicarb (Temik®) or imidacloprid (Admire®). These insecticides are aimed at controlling psyllids, a possible insect vector for the disease, although it is not known if insecticides have a direct effect on the spread of HLB. These insecticide treatments do not reduce disease in trees already infected, in any case. An integrated control program has been recommended for HLB in commercial orchards by the United Nations Development Program, Food and Agriculture Organization (UNDP, FAO) Southeastern Asian citrus rehabilitation project (Aubert, 1990). The program highlights controlling psyllid vectors with insecticides, reducing inoculum through removal of HLB-symptomatic trees, propagating and using pathogen-free budwood and nursery trees. In Florida, foliar nutrition programs coupled with vector control are often used to slow down the spread of HLB and reduce devastating effects of the disease (Gottwald, 2010). These control practices have shown limited effect for preventing the further spread of HLB. Other than destruction and removal of diseased trees, there is no effective control for HLB in infected trees, and there is no known cure for HLB. New and improved treatments for citrus (and other) HLB disease therefore are needed in the art.
Other plant pathogens of the greatest interest include the bacterium Xanthomonas citri causing citrus canker, Xanthomonas axonopodis pv. citrumelo causing citrus bacterial spot disease, and Xylella fastidiosa causing citrus variegated chlorosis; the pathogenic fungus Alternaria citri causing leaf and stem rot and spot, Phytophthora spp. causing foot and root rot, and Guignardia citricarpa causing citrus black spot, all of which can result in crop loss.
Induced resistance can confer long-lasting protection against a broad spectrum of plant diseases either locally or systemically (Durrant and Dong, 2004; Walters et al., 2013). Plant defense mechanisms can be activated by pathogens (Durrant and Dong, 2004), beneficial microorganisms (Weller et al., 2012; Zamioudis and Pieterse, 2012), or by chemical inducers (Walters et al., 2013). Overall, maximizing crop plant health and vigor has been a difficult problem with no comprehensive solution. Therefore, the embodiments of the invention described herein are provided for the control of crop pathogens such as HLB, Xanthomonas citri causing citrus canker, Xanthomonas axonopodis pv. citrumelo causing citrus bacterial spot disease, and Xylella fastidiosa causing citrus variegated chlorosis; the pathogenic fungus Alternaria citri causing leaf and stem rot and spot, Phytophthora spp. causing foot and root rot, and Guignardia citricarpa causing citrus black spot and to improve plant health and vigor, including germination, growth, disease resistance, and improvement of crop quality and quantity.